The immune system has the ability to mount responses that can destroy tumor cells. Among the various elements of the immune system, cytotoxic T lymphocytes (CTL) are highly effective in mediating the rejection of established tumors. CTL's recognize antigenic determinants produced from any protein synthesized within the cell, while antibodies recognize and bind only integral cell surface molecules.
The anti-tumor activity of tumor-specific CTL is the result of a series of complex molecular events. After cellular processing of proteins in the cytoplasm of the tumor cells, small peptides are transported to the endoplasmic reticulum, where they bind to newly synthesized major histocompatibility gene complex (MHC) class I molecules or HLA's. HLA/peptide complexes are then exported to the surface of the tumor cell where they are recognized by antigen-specific Class I-restricted CTL's. In addition to lysing the tumor cell, the CTL's may also secrete lymphokines such as tumor necrosis factor (TNF), and gamma-interferon (γ-IFN), which also contribute to the overall anti-tumor effect.
A family of genes referred to as the “MAGE” family after the melanoma associated antigen encoding gene, MAGE-1, has been discovered which are processed into peptides and expressed on tumor cell surfaces as HLA/peptide complexes. The MAGE peptides are recognized by specific CTL's leading to lysis of the tumor cells from which they are expressed. The genes code for “tumor rejection antigen precursors” and the peptides derived therefrom are referred to as “tumor rejection antigens” (see Traversari et al. (1992) Immunogenetics 35:145; van der Bruggen et al (1991), Science 254:1643; and U.S. Pat. No. 5,342,774; all of which are herein incorporated by reference).
In fact, the MAGE-1 encoded human melanoma specific antigen, MZ2-E, is recognized by CTL's derived from a cancer patient (Van der Bruggen et al. (1991) Science 254:1643–1647). The MAGE-1 gene is expressed by various melanoma cell lines as well as several other types of tumor cells, but is not expressed in a panel of normal tissues. Eleven additional members of the MAGE family, map to the q28 region of chromosome X and have between 64% and 85% identity in amino acid sequence to MAGE-1 (Chen et al. (1994) Proc. Natl. Acad. Sci. 91:1004–1008); De Plaen et al. (1994) Immunogenetics 40:360–369; Wang et al. (1994) Cytogenet. Cell Genet. 67:116–119). These genes on the q28 region of chromosome X are referred to as the MAGE-A family genes (MAGE-A1 to A12) (Duffour et al. (1999) Eur. J. Immunol. 29:3329–3337).
The MAGE-A family of genes have been found to be expressed at a high level in a number of tumors of various histologic types including those from colorectal, lung, ovarian, breast, colon, lung, liver, thyroid, and skin cancers (Mori et al. (1996) Ann. Surg. 183–188; Sakata M. (1996) Kurume Med. J. 43:55–61; Yamada et al. (1995) Int. J. Cancer 64:388–393; Zukut R. et al. (1993) Cancer Res. 53:5–8; Zakut R. et al. (1990) Cancer Res. 53:5–8). In addition, significantly increased levels of MAGE-A4 have been detected in the sera of patients with hepatitis C virus (HCV)-associated cancer and HCV-associated liver cirrhosis indicating (Tsuzurahara et al. (1997) Jpn. J. Cancer Res. 88:915–918.) Examination of a large panel of healthy tissues revealed expression of MAGE genes only in testis and placenta (De Plaen et al., supra).
For example, the MAGE-A1 gene is expressed in approximately 40% of melanomas and in some other tumors and the MAGE-A2 and MAGE-A3 genes are expressed in approximately 80–90% of the melanoma lines that have been examined. Activation of MAGE-A1 in cancer cells may be due to demethylation of the promoter sequence. Treatment with the demethylating agent 5-aza-2′-deoxycytidine activated MAGE-A1 expression not only in tumor cell lines, but also in primary fibroblasts (De Smet C. et al. (1996) Proc. Natl. Acad. Sci. 93:7149–7153).
Another family of tumor rejection antigen precursor genes has been identified on the Xp arm of the X chromosome. These genes are referred to as the MAGE-B family of genes (MAGE-B 1 to B4). The MAGE-B 1 and MAGE-B2 genes are similarly expressed in tumors of various histological origins, silent in normal tissues with the exception of testis, and activated by a demethylation process (Lurquin et al. (1997) Genomics 46:397–408). In addition, studies by McCurdy et al. indicate that MAGE Xp-2 (MAGE-B2) is the target of autoantibodies in systemic Lupus Erythematosus (SLE), suggesting that this protein may also have a role in autoimmune and inflammatory disorders (McCurdy et al. (1998) Molec. Genet. Metab. 63:3–13). A gene designated MAGE-C1 has also been identified.
The identification of tumor specific antigens, such as those encoded by MAGE genes, and the corresponding T cell epitopes have provided novel peptide-based vaccines useful in treating cancer patients. For example, a nonapeptide fragment of MAGE-A1 stimulates CTL's that respond to antigen MZ2-E (Traversari et al. (1992) J. Exp. Med. 176:1453–1457). Cells that present the nonapeptide, EADPT-GHSY, were used to immunize MAGE-A positive melanoma patients (Hu et al. (1996) Cancer Res. 56:2479–2483). The immunization increased the frequency of autologous melanoma-reactive CTL precursors in the circulation. In combination with interleukin-2 the MAGE-A1 nonapeptide immunization led to a significant expansion of the peptide-specific and autologous melanoma-reactive CTL response (Hu et al., supra). In addition, a tumor rejection antigen derived from MAGE-A3 tumor rejection antigen precursors is presented by HLA-A1 molecules, and tumor rejection antigens derived from MAGE-A2 complex with MHC class I molecule HLA-A2.
More recently it has been demonstrated that an anti-MAGE-A4 tumor rejection antigen CTL clone can lyse HLA-A2 tumor cells expressing MAGE-A4 tumor rejection antigen precursors (Duffour et al., supra). These data are especially important as MAGE-A4 is expressed in 51% of lung carcinomas and 63% of esophageal carcinomas, whereas about 50% of Caucasians and Asians express HLA-A2. These results indicate that MAGE proteins other than MAGE-A1 are likely to be valuable for cancer immunotherapy.
In addition to their value as anticancer agents, there is evidence that MAGE proteins may also have use in the treatment of neurodegenerative conditions. For example, MAGE genes are related to the necdin gene, and the small potential transmembrane domain of the MAGE proteins shows a particularly high degree of conservation with the transmembrane domain of the necdin protein. It has been postulated that this region associates with the transmembrane domain of another protein (De Plaen et al, supra).
Necdin is a nuclear protein, first identified in neuronally differentiated embryonal carcinoma cells and in the brain of adult mice (Maruyama et al. (1991) Biochem. Biophys. Res. Commun. 178:291–296). Necdin is expressed in virtually all postmitotic neurons in the central nervous system at all stages of development (Uetsuki et al. (1996), J. Biol. Chem. 271:918–924). However, necdin is not expressed in proliferative neuron-like cells originating from tumors, and ectopic expression of necdin in NIH3T3 cells suppresses cell growth without affecting cell viability (Aizawa et al., (1992) Dev. Brain Res. 63:265–274); Hayashi et al., (1995) Biochem. Biophys. Res. Commun. 213:317–324). Therefore, necdin is likely to affect the transition in developing neurons from proliferative to non-proliferative states (Uetsuki et al., supra). Furthermore, necdin has been shown to interact with viral transforming proteins such as SV40 large T antigen and adenovirus E1A, and with the transcription factor E2F1. Necdin can also functionally replace the Rb as a growth suppressor in Rb deficient osteosarcoma cells, suggesting that necdin is a neuron-specific growth suppressor with a function similar to that of Rb (Taniura et al. (1998) J. Biol. Chem. 273:720–728). Therefore, MAGE proteins may also function to suppress growth in neuronal cells and, thus, be involved in the pathophysiology of neurodegenerative conditions.
It is well established that members of the MAGE protein family play critical roles in a variety of important cellular processes including the regulation of cellular growth and differentiation; T-cell activation; CTL effector cell function; and elicitation of auto-antibodies. As a result of these roles, the MAGE proteins are involved in such important diseases and disorders as cancers, tissue repair, neurodegenerative disorders, autoimmune disorders, and inflammatory disorders.
Accordingly, the discovery of polynucleotides encoding MAGE-like proteins, and the proteins themselves, provides a means to investigate MAGE-mediated disorders, and provides new compositions useful in the diagnosis and/or treatment of cancers, neurodegenerative disorders, autoimmune disorders such as SLE, and inflammatory disorders.